In the year 2000, an estimated 22 million people were suffering from cancer worldwide and 6.2 millions deaths were attributed to this class of diseases. Every year, there are over 10 million new cases and this estimate is expected to grow by 50% over the next 15 years (WHO, World Cancer Report. Bernard W. Stewart and Paul Kleihues, eds. IARC Press, Lyon, 2003). Current cancer treatments are limited to invasive surgery, radiation therapy and chemotherapy, all of which cause either potentially severe side-effects, non-specific toxicity and/or traumatizing changes to ones body image and/or quality of life. Cancer can become refractory to chemotherapy reducing further treatment options and likelihood of success. The prognosis for some cancer is worse than for others and some are almost always fatal. In addition, some cancers with a relatively high treatment success rate remain major killers due to their high incidence rates.
One of the causes for the inadequacy of current cancer treatments is their lack of selectivity for affected tissues and cells. Surgical resection always involves the removal of apparently normal tissue as a “safety margin” which can increase morbidity and risk of complications. It also always removes some of the healthy tissue that may be interspersed with tumor cells and that could potentially maintain or restore the function of the affected organ or tissue. Radiation and chemotherapy will kill or damage many normal cells due to their non-specific mode of action. This can result in serious side-effects such as severe nausea, weight loss and reduced stamina, loss of hair etc., as well as increasing the risk of developing secondary cancer later in life. Treatment with greater selectivity for cancer cells would leave normal cells unharmed thus improving outcome, side-effect profile and quality of life.
The selectivity of cancer treatment can be improved by using antibodies that are specific for molecules present only or mostly on cancer cells. Such antibodies can be used to modulate the immune system and enhance the recognition and destruction of the cancer by the patient's own immune system. They can also block or alter the function of the target molecule and, thus, of the cancer cells. They can also be used to target drugs, genes, toxins or other medically relevant molecules to the cancer cells. Such antibody-drug complexes are usually referred to as immunotoxins or immunoconjugates and a number of such compounds have been tested in recent year [Kreitman R J (1999) Immunotoxins in cancer therapy. Curr Opin Immunol 11:570-578; Kreitman R J (2000) Immunotoxins. Expert Opin Pharmacother 1:1117-1129; Wahl R L (1994) Experimental radioimmunotherapy. A brief overview. Cancer 73:989-992; Grossbard M L, Fidias P (1995) Prospects for immunotoxin therapy of non-Hodgkin's lymphoma. Clin Immunol Immunopathol 76:107-114; Jurcic J G, Caron P C, Scheinberg D A (1995) Monoclonal antibody therapy of leukemia and lymphoma. Adv Pharmacol 33:287-314; Lewis J P, DeNardo G L, DeNardo S J (1995) Radioimmunotherapy of lymphoma: a UC Davis experience. Hybridoma 14:115-120; Uckun F M, Reaman G H (1995) Immunotoxins for treatment of leukemia and lymphoma. Leuk Lymphoma 18:195-201; Kreitman R J, Wilson W H, Bergeron K, Raggio M, Stetler-Stevenson M, FitzGerald D J, Pastan I (2001) Efficacy of the anti-CD22 recombinant immunotoxin BL22 in chemotherapy-resistant hairy-cell leukemia. N Engl J Med 345:241-247]. Most antibodies tested to date have been raised against known cancer markers in the form of mouse monoclonal antibodies, sometimes “humanized” through molecular engineering. Unfortunately, their targets are usually also present on subset of normal cells thus still causing some non-specific effect. Furthermore, these antibodies are basically mouse proteins that are being seen by the human patient's immune system as foreign proteins. The ensuing immune reaction and antibody response can result in a loss of efficacy or in side-effects.
The inventors have used a different approach in their development of antibodies for cancer treatment. Instead of immunizing experimental animals with cancer cells or isolated cancer cell markers, they have sought out only those markers that are recognized by the patient's own immune system or, in other words, that are seen by the immune system as a foreign molecule. This implies that the markers or antigens are usually substantially absent on normal cells and, thus, the risk of non-specific toxicity is further reduced. Hybridoma libraries are generated from cancer patient-derived lymphocytes and the antibodies they secrete are tested for binding to normal and tumor cells. Only antibodies showing high selectivity for cancer cells are retained for further evaluation and development as a cancer therapeutic or diagnostic agent. One such highly selective antibody is the subject of this patent application. In addition to being selective, this antibody is fully compatible with the patient's immune system by virtue of being a fully-human protein. The antibody of the invention can be used for diagnostic or therapeutic uses or as a basis for engineering other binding molecules for the target antigen. The antibody of the invention can also be used to identify the target antigen. The antigen can then be used to design new cancer treatment or diagnostics.
The basic structure of an antibody molecule consists of four protein chains, two heavy chains and two light chains. These chains are inter-connected by disulfide bonds. Each light chain is comprised of a light chain variable region and a light chain constant region. Each heavy chain is comprised of a heavy chain variable region and a heavy chain constant region. The light chain and heavy chain variable regions can be further subdivided into framework regions and regions of hypervariability, termed complementarity determining regions (CDR). Each light chain and heavy chain variable region is composed of three CDRs and four framework regions.
Glucose transporter 8 (GLUT8) is a member of the GLUT family of proteins and is known to have sugar transporting activity. GLUT8 is encoded by gene slc2a8, which is found on human chromosome 9. GLUT8 is 477 amino acids in length. It is a ˜50 kDa type II transmembrane protein. It has 12 transmembrane regions. It has a short extracellular loop between TM1 and TM2 and a long extracellular loop between TM9 and TM10. Despite having several transmembrane regions, GLUT8 is located intracellularly likely because of a N-terminal di-leucine motif (Ibberson et al. JBC 275: 4607-4612, 2000; Moadel et al., Cancer Res 65: 698-702, 2005). Translocation to the membrane has been observed in mouse cells upon insulin treatment (Carayannopoulos et al., PNAS 97:7313-18, 2000) or in rat cells upon hypoxic shock or insulin treatment (U.S. Ser. No. 09/886,954 [2002/0038464]). In human, membrane localization has not been reported and no stimuli has been identified to induce translocation (Widmer et al., Endocrinology 146:4727-36, 2005).
GLUT/SLC2A family nomenclature has been published in: Amer. J. Physiol. Endocrinol. Metab. 282:E974-76, 2002. The name GLUT8 was used in the past to describe what it now known as GLUT12—as indicated in that paper. The N-terminal di-leucine motif has been found in all mammalian GLUT8 sequences (see Zhao et al., Biochimica et Biophysica Acta 1680:103-113, 2004—showing bovine, human, rat, mouse).